Many compounds are known to affect neuromuscular transmission. It is known that some of these compounds block neuromuscular transmission by blocking the channel activated by acetylcholine, either in its open or closed conformation. The objective of this proposal is to determine the mechanism by which guanidine derivatives block the closed acetylcholine-activated channels of frog and tissue-cultured muscle. The actions of these compounds on nerve or iontophoretically-induced endplate currents will be determined at the voltage endplates of cutaneous pectoris muscles. The effects of the guanidine derivatives on single channel currents in tissue cultured cells will be examined using the patch clamp technique. The ability of a number of structurally related guanidien derivatives to block the closed channel will be determined and a staructure-activity relationship foarmulated. The voltage, time and concentration dependence of closed channel block will be examined. Dose-response relationships for the ability of the derivatives to block the closed channel will be constructed using the known voltgage, time and concentration dependence. Measurements of single channel paroperties bill be made to examine the action of these compounds on single channel conductance, channel open time and channel closed time. This information will provide information on whether or not population effects or reduction in single channel current amplitude are involved in closed channel block. This information will be used to test the validity of several atheories regarding the mechanism of closed channel block and to propose a resonable mechanism by which this action can occur. The data gathered during this project should help us to better understand other drug actions (i.e., open channel block), shed light on the mechanism of ion permeation and provide us with information regarding channel structure.